核糖体基因间间隔区分析
DNA提取
标准化
土壤水分
土工试验
萃取(化学)
再现性
环境科学
微生物种群生物学
土壤质量
生物
土壤科学
核糖体RNA
聚合酶链反应
色谱法
计算机科学
化学
内转录区
基因
遗传学
细菌
操作系统
作者
Imes Petric,Laurent Philippot,Cristina Abbate,Antonio Bispo,Thierry Chesnot,Sara Hallin,Karine Laval,Thierry Lebeau,Philippe Lemanceau,Corinne Leyval,Kristina Lindström,Pascal Pandard,Esperanza Romero,Amadou Sarr,Michael Schloter,Pascal Simonet,Kornelia Smalla,Berndt‐Michael Wilke,Fabrice Martin-Laurent
标识
DOI:10.1016/j.mimet.2011.01.016
摘要
Extracting DNA directly from micro-organisms living in soil is a crucial step for the molecular analysis of soil microbial communities. However, the use of a plethora of different soil DNA extraction protocols, each with its own bias, makes accurate data comparison difficult. To overcome this problem, a method for soil DNA extraction was proposed to the International Organization for Standardization (ISO) in 2006. This method was evaluated by 13 independent European laboratories actively participating in national and international ring tests. The reproducibility of the standardized method for molecular analyses was evaluated by comparing the amount of DNA extracted, as well as the abundance and genetic structure of the total bacterial community in the DNA extracted from 12 different soils by the 13 laboratories. High quality DNA was successfully extracted from all 12 soils, despite different physical and chemical characteristics and a range of origins from arable soils, through forests to industrial sites. Quantification of the 16S rRNA gene abundances by real time PCR and analysis of the total bacterial community structure by automated ribosomal intergenic spacer analysis (A-RISA) showed acceptable to good levels of reproducibility. Based on the results of both ring-tests, the method was unanimously approved by the ISO as an international standard method and the normative protocol will now be disseminated within the scientific community. Standardization of a soil DNA extraction method will improve data comparison, facilitating our understanding of soil microbial diversity and soil quality monitoring.
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